Summary One in three HIV-infected individuals develops some form of HIV-associated neurocognitive disorder (HAND). Consumption of drugs of abuse such as methamphetamine (METH) aggravates the symptoms of HAND, but the cellular and molecular mechanisms by which these drugs impact HIV disease progression in the central nervous system (CNS) remain ill-defined. In this study we will thoroughly test the hypothesis that HAND is the result of the neurotoxic effects of HIV proteins synthesized when latently infected microglial cells respond to signs of neurodegeneration. We will also test the hypothesis that exposure to drugs of abuse enhance HIV replication and exacerbate HAND. This highly multidisciplinary investigation is a close collaboration between the laboratories of Dr. Jonathan Karn (CWRU), an expert in the molecular mechanisms HIV latency, and Dr. Kurt Hauser, a neurobiologist and expert on drug abuse (VCU). We have recently established a reliable and robust method to develop immortalized microglial cells from primary glia derived from fresh CNS tissue, and used them to create microglia/HIV cellular models. The proposal capitalizes on findings of an unbiased shRNA library screen, which revealed that the Nurr1/CoREST trans- repressor complex plays a key role in silencing HIV in microglial cells, a mechanism which is distinct from silencing in memory T-cells. Building on these observations, we will study the epigenetic machinery leading to silencing of the HIV promoter, including Nurr1/CoREST, by chromatin immunoprecipitation experiments (Chip-Seq), and study the impact of both acute and chronic treatment with METH on reversing HIV latency. Our experiments will also uncover the inflammatory signals that, together with METH, induce HIV expression. Finally, using two novel co-culture systems, we will demonstrate how METH-primed microglia/HIV cells exacerbate neuronal damage. Successful completion of these studies will provide a detailed understanding of fundamental biology of HIV-infected microglia in response to METH. By establishing the relationship between HIV latency, inflammation, and neuronal damage we will provide new insights into the development of HAND in HIV patients and how this is augmented by METH abuse.